Natural Killer cells play a significant role in host defense, in part by attacking transformed, infected or normal cells that downregulate MHC class I molecules. Target cell recognition by NK cells involves both stimulatory and inhibitory receptors, the latter of which are specific for class I molecules. Inhibitory receptors are expressed by overlapping subsets of NK cells in a pattern that appears largely random. The loss of inhibitory signaling by MHC-specific receptors is responsible for the capacity of NK cells to attack class I-deficient cells. Because inhibitory receptors discriminate allelic variants of class I molecules, NK cells also attack MHC-different cells. Therefore, a key issue is how NK cells are rendered tolerant to self cells. The leading hypothesis, cited even in textbooks, is that each NK cell is endowed with at least one inhibitory receptor specific for a self MHC class I molecule. In the last funding period, we have developed evidence that this is not the case. A fraction of NK cells in normal mice lacks self MHC-specific receptors, yet is self-tolerant. These NK cells fail to attack class I deficient lymphoblasts or bone marrow cells. Importantly, these cells are hyporesponsive to some other defined stimuli, such as crosslinking of NKG2D or NKR-P1 stimulatory receptors. NK cells that arise in class I-deficient mice exhibit a similar hyporesponsive phenotype. Our hypothesis is that inhibitory receptor expression, being substantially stochastic during NK development, fails to endow a fraction of NK cells with self MHC-specific receptors. These NK cells achieve self-tolerance by adopting a hyporesponsive phenotype that precludes them from attacking self cells, yet allows them to function in other contexts. All the NK cells in class I-deficient mice adopt this hyporesponsive phenotype, because they fail to encounter inhibitory MHC molecules during development. To test this hypothesis and address the underlying molecular mechanisms, we propose the following specific aims: Specific Aim 1: To identify genes whose expression levels differ in responsive versus hyporesponsive NK cell populations. Specific Aim 2: To determine the signaling pathways and the steps in the pathway that are impaired in hyporesponsive NK cells. Specific Aim 3: To determine the relationship of NK cell hyporesponsiveness and cytokine action.